At the neuromuscular junctions (NMJs) of aged CBF-1 mice, an adaptive change (increase in transmitter release) apparently permits synaptic function to be well-preserved despite the presence of fewer synaptic vesicles and fewer mitochondria. This proposal is a step in elucidating the cellular mechanisms of these compensations. Hypotheses about this adaptation will be tested: 1) The number, length, and particle organization, of presynaptic active zones and spatial distribution of synaptic vesicles, both of which critical factors in transmitter release, may change with age; 2) Synaptic vesicles may be recycles faster at the aging NMJ; 3) Altered calcium entry at the axon terminal or calcium ion balance in the pre-terminal axon may underlie the adaptive changes 4) Furthermore, it is important to determine whether the structure change proceeds, accompanies, or follows the functional change. Differences between aged and young male CBF-1 mouse NMJ will be evaluated in the following ways: 1) Serial and semi- serial sections of end plates will be examined to determine the number and length of active zones and the spatial distribution of synaptic and coated vesicles in synaptic and non-synaptic areas. 2) Freeze fracture of nerve terminals active zones. 3) Morphological analysis of synaptic vesicle recycling will be carried out by applying long term tetanic stimulation in combination with HRP and cationized Ferritin, in order to quantify and assess the extent of vesicle replacement. 4) Changes in the relationship of evoked transmitter output to different extracellular calcium-magnesium concentrations and analyze the time course of intracellular calcium sequestering on spontaneous transmitter release. 5) The progression of age-related changes in neuromuscular structure will be investigated using scanning and transmission electron microscopy procedures in conjunction with light microscopy of the nerve terminal arborization. The long term objectives of this proposal are a) to characterize the time- course of change and causal factors that underlie synaptic plasticity during aging, and b) to provide a basis for attempting to pharmacologically enhance synaptic function in the aging motor system.